In a unique bi-phasic approach, cell surface glycosphingolipids (GSL) of human lymphocytes will be examined. First, by the use of high-performance liquid chromatography (HPLC), quantitative analysis of as few as 5 times 10 to the 6th power normal or transformed human lymphocytes can yield precise quantitative data on the GSL content of these cells. Secondly, antibodies to purified, chemically characterized, GSL will be used in immunofluorescent studies of surface properties of lymphocyte GSL antigens. These antibodies can be rendered specific for a defined carbohydrate sequence by immunoadsorption, in most cases reactive only with the immunizing antigen. The highly restricted specificity of these antibodies will be used to explore subpopulations of normal and transformed human lymphocytes. Together with the chemical data, the immunochemical techniques will be employed in distribution and functional studies and to probe the mechanism of membrane alterations associated with transformation. Many changes occur in GSL content of transformed cells, and these changes are thought to play a role in altered growth patterns of these cells. Human lymphocytes have received little attention in this regard, while changes in antigenicity of GSL in transformed cells has not been studied at all. I propose to integrate studies of chemical and antigenic properties of normal and transformed human lymphocytes, in order to explore the relationship between the mechanism of transformation and the apparent alterations in GSL content.